EXTRACŢIA CAPSAICINEI DIN ARDEIUL IUTE

Generalităţi
Capsaicina se extrage în principal din ardeiul iute.

Parte experimentală
Capsaicina se extrage din ardeiul iute folosind solvenţi organici (alcool acetona,
acetonitril, hexan) şi metode diferite de extracţie.

1. Extracţia capsaicinei prin macerarea ardeiului iute în alcool.

Ardeiul se mărunţeşte şi se adaugă într-un orcan închis ermetic. !este ardei se introduce
alcool etilic concentrat (sau vodca) p"nă ce planta este complet imersată în alcool. #e menţine, la
soare cel puţin $% ore, după care se filtrea&ă se concentrea&ă prin evaporarea parţială a alcoolului
şi se introduce într-un recipient închis ermetic. #e adaugă puţin amestec la o cremă de corp şi se
oţine o cremă împotriva durerilor musculare.

2. Extracţia capsaicinei cu acetonă

!atru 'ilograme ardei iute se mărunţesc în ender fără a adăuga nici un fel de lichid.
Amestecul oţinut este uscat prin liofili&are şi menţinut la -*C.
#e c"ntăresc + g material uscat şi se extrag, timp de  h cu +, l acetonă, la rece. upă
 h se filtrea&ă lichidul. !rocesul se repetă p"nă soluţia extrasă este incoloră (cca -$ ori). /inal
materialul extras răm"ne su formă de pulere fină de culoare uşor găluie. #oluţia de acetonă
(de culoare verde închis datorită extracţiei clorofilei) sunt amestecate şi concentrate la
rotavapori&ator. Amestecul concentrat rămas (ce conţine oleorăşini) este diluat cu apă deioni&ată
luată de $ ori volumul soluţiei concentrate, apoi adusă la p01++ cu sol. ,2 340. Amestecul se
diluea&ă în continuare cu +, volume apă deioni&ată, după care se aduce la p015, cu soluţie
0Cl şi se extrage de  ori cu eter etilic într-o p"lnie de separare. 6terul se distilă la vid şi î la& se
recoltea&ă un ulei v"scos de culoare run-ver&ui.

3. Extracţia capsaicinei prin metoda Soxhlet 

Ardeiul se taie în sferturi, se separă manual seminţele (acestea au cel mai mare conţinut
în capsaicină) şi se taie c"t mai fin sau se mărunţesc la ender. #e usucă materialul vegetal în
etuvă la temperaturi su +*C.
#e montea&ă alonul pe o plită de încăl&ire sau pe o aie de apă şi se adaugă + ml
etanol 78. #e montea&ă instalaţia #oxhlet şi se începe încăl&irea. 9efluxarea se continuă p"nă
c"nd etanolul ce se adună în piesa intermediară este incolor. 6xtractul se concentrea&ă la
rotavapori&ator sau distilare la vin a etanolului. Ametecul concentrat se menţine peste noapte
într-un flacon închis ermetic.
 

Atenţie!!!!
Capsaicina pură provoacă arsuri ale pielii în cantităţi foarte mici. :a fel şi oleorăşina
extrasă din ardei.
C"nd lucraţi folosiţi echipamentul complet de protecţie; ochelari, mănuşi, halat, şi, dacă e
 posiil, şi mască împotriva prafului.

1. If available, use a rotary evaporator with a bump-trap to remove solvent.

2. Set up a distillation apparatus, evaporate solvent under reduced pressure carefully to prevent
bumping
3. Allow concentrate to stand overnight in a sealed vial once cool. il will form separate layer.
!. "se pasteur pipette to remove bottom oil layer #red-brown$.

%haracteri&ation of Scoville 'ating(

• A panel of ) volunteers tasted 1(1* serial dilutions of the original concentrate in )+ sucrose.
• Spicy tastesensation was noted at the 1(1** dilution by !), and again at 1(1* with !) #different
individual as the negative result individual$
• %oncentrate was tasted after concluded to be safe enough to try  volunteers noted that the
concentrate was comparable to tobasco sauce.

uture improvements(
• Avoid the vacuum distillation process, as some capsaicinoids could have been lost at this step
• btain better peppers, dry fully, and grind up well #this step was bypassed in lieu of filming the
video /uic0ly$
• erform % to determine composition of dilute e4tract and concentrate.

Abstract

 A new method based on pressurized liquid extraction followed by LC-MS/MS analysis has been
developed for the identification and quantification of three capsaicinoids capsaicin! dihydrocapsaicin! and
nordihydrocapsaicin" in extracts of Capsicum annuum# $or the recovery of three capsaicinoids! the
efficiency levels of ultrasonic-assisted extraction! microwave-assisted extraction! Soxhlet extraction! and
pressurized liquid extraction were compared under different conditions# %ressurized liquid extraction
resulted in hi&her yields# %ressurized liquid extractions were performed usin& methanol' temperature was
set at ())*C and pressure at (+)) psi# LC analysis was performed on a ,aters .rid&e C(0 column
(+) 1 2#( mm! id 3#+ 4m" eluted by a mobile phase of )#(5 formic acid and AC6# 7ata acquisition was
carried out in multiple reaction monitorin& transitions mode! monitorin& two-reaction monitorin& transitions
to ensure an accurate identification of tar&et compounds in the samples# 8he proposed method is rapid!
simple! and could be utilized for the routine analysis of three capsaicinoids in C# annuum samples#
 

A simple method for the analysis of capsaicin and dihydrocapsaicin in peppers by ultrasound assisted e4traction #"SA5$
followed by gas chromatographymass spectrometry #6%-7S$ has been developed. "SA5 conditions were optimi&ed by
e4perimental design in order to ma4imi&e analyte e4traction. A full factorial design involving e4traction variables such as
solvent #ethanol and water$, e4traction time #)2) min$, e4traction temperature #2))* 8%$, sample amount #*.2)*.) g$,
and ultrasound amplitude #!*9*+$ was applied. he most significant conditions for capsaicinoid e4traction by "SA5 were
solvent type, e4traction time, and sample amount. he obtained results were compared with other e4traction methods( the
official So4hlet method and a previously reported solid phase microe4traction method. 'esults showed that the e4traction
efficiency with the application of "SA5 #:9+$ was as good as that obtained with So4hlet and the precision of recovery was
less than )+; in addition, the e4traction time was decreased from ) h to 2) min. he 6%-7S analytical method was linear
in the range 1*1** <gm for capsaicin and dihydrocpsaicin with correlation coefficient r = *.::9 and pea0 area
variability of 1+ for both capsaicinoids. he method was applied to the analysis of 11 varieties of hot peppers cultivated
in 7>4ico. A large concentration range for capsaicin #1*1?9** <gg$ and dihydrocapsaicin #11*2@3? <gg$ was found in
these pepper samples.

Chili peppers are the fruits of the genus Capsicum. There are several chili pepper varieties and they have
 been domesticated for years. Chili peppers are used to season foods in many cultures and they have
 become a staple in the diets of Italians, Spaniards, Hungarians, Indians, Chinese, Indonesians, and
 Africans.
The chemicals that cause the sensation of heat when you eat a pepper are called capsaicinoids. Capsaicin
 being the main capsaicinoid found in chili peppers. This chemical is so hot that just one drop mied with
!",""" drops of water will cause the tongue to blister.
 #hen you eat a pepper the capsaicin irritates the lining of your mouth causing that familiar burning
sensation. $our brain responds to this stimulus by producing endorphins, your body%s own natural pain
reliever. The production of endorphins in response to eating chili peppers may eplain why they can be
addicting.
The capsaicin found in peppers also has medicinal uses. It relaes the arteries &which lowers blood
pressure' and can be used in creams as a topical pain reliever. Currently, capsaicin and capsaicin
derivatives are used to treat arthritis, shingles, diabetic neuropathy, neuralgia, and as a pain reducer
following surgery. Some studies have even shown that consumption of capsaicin(containing peppers
reduces the amounts of clotting protein in you blood, thereby reducing a person%s ris) of having a stro)e.
The hotness of a chili pepper is measured in Scoville units. These units were invented by  Wilbur
Scoville, a pharmacist, in !*!+. To determine the hotness- using Scoville units, a panel of  tasters
sample etracts of the peppers that are diluted in sugar water. The dilution that can be detected by / of the
 tasters is considered the heat level of the pepper.
The ratio of the etract dilution is the Scoville unit. 0or eample, the jalapeno is usually detected by / of 
 tasters when diluted at a ratio of !1+"" to !1"""2 the Scoville unit for the jalapeno is +"" to """.
3ust to compare, the habanero pepper can have !"",""" to /"",""" Scoville units.
In this lab, you will be using a variation of Scoville%s test to determine the hotness of several types of 
capsicum peppers.

Materials
glass mortar 4 pestle  +(dram vials with caps graduated pipettes late gloves
5 sucrose solution *5 ethanol electronic balance
 weighing boat )nife various chili peppers

Procedure
!. 6et into a group of  an obtain the necessary materials. 7ach person in your group should obtain
a different type of pepper.
+. 8abel the dram vials !9!", !9 !"", !9 !,""", !9 !",""", !9!"",""".
/. 0ill each dram vial with :. m8 of 5 sucrose solution.
& If you are unsure about how to read the pipette, please ask.'
:. ;lace the weighing boat on the electronic balance. <ero the balance by pushing the =ero- button.
. Cut off a small piece of the pepper and place it in the weighing boat. $ou want a  grams sample of 
 

pepper. Cut off or add more pepper until the balance reads  grams &re(=ero the balance if 
necessary'.
>. ;lace the pepper sample in the glass mortar and add m8 of *5 ethanol. ?se the pestle to grind
up the pepper in the ethanol.
@. ?se a clean pipette to transfer ".m8 of the etract into the vial mar)ed !9!". Cap and sha)e
the vial.
. ?se a clean pipette to transfer ".m8 of the etract in the !9!" vial into the vial mar)ed !9!"".
Cap and sha)e the vial.
*. ?se a clean pipette to transfer ".m8 of the etract in the !9!"" vial into the vial mar)ed
!9!,""". Cap and sha)e the vial.
!". ?se a clean pipette to transfer ".m8 of the etract in the !9!,""" vial into the vial mar)ed
!9!",""". Cap and sha)e the vial.
!!. ?se a clean pipette to transfer ".m8 of the etract in the !9!",""" vial into the vial mar)ed
!9!"",""". Cap and sha)e the vial.
!+. ;lace a clean pipette in the !9!"",""" vial.
!/. ;erform the taste test with the other four members of your group. ?sing the clean pipette in the
!9!"",""" vial to place a drop of the solution on each person%s coffee stirrer. All members should
place the drop on their tongues and determine if they can detect the capsaicin.
!:. Bove the same pipette &you do not need a new one for each vial' to the net vial &!9!","""' and
repeat the taste test. Continue until at least three people in your group can detect the capsaicin in
the solution, then mar) the appropriate bo in the data table.
!. 7at an unsalted crac)er and drin) some water.
!>. epeat steps !+(! until all  peppers have been tested.
!@. ecord the Scoville units in the data table. emember that if capsaicin is detected at the !9!","""
dilution, then the Scoville unit for that pepper is !",""".
!. #ash and dry all of the materials and place them bac) in the proper place. $our bo of materials
should have the mortar 4 pestle, !" dram vials, and ! pipettes in it before you leave the room.

AOAC Offiial et"#$ %%&'()

Cap*aiin#i$* in Cap*i+m* an$ T"eir Extrati,e*
Li-+i$ C"r#mat#.rap"i et"#$
/ir*t Ati#n 0%%&
1Applia2le f#r $eterminati#n #f 3&(45&(((( S#,ille 6eat Unit* 7S6U8 #f 
ap*aiin#i$* in .r#+n$ an$ r+*"e$ re$ pepper9 "ili pepper9 .r#+n$ a:enne
 

pepper9 .r#+n$ ;alapen# pepper9 an$ re$ pepper #le#re*in*' N#t applia2le t# "ili
p#<$er* #r pr#$+t* #ntainin. #re.an# #r t":me'=
(Caution: See Appendix B9 *afet: n#te* f#r t"e *afe "an$lin. #f #r.ani *#l,ent* an$
*peial "emial "a>ar$*4aet#ne9 aet#nitrile9 an$ et"an#l' See aterial Safet:
Data S"eet*9 #r e-+i,alent9 f#r ea" rea.ent' N4?anill:i4nn#nanami$e i* an extreme
irritant@ $# n#t in"ale' Di*p#*e #f <a*te *#l,ent* in an appr#priate manner
#mpati2le <it" applia2le en,ir#nmental r+le* an$ re.+lati#n*'8
et"#$ Perf#rmane
See Ta2le %%&'() f#r met"#$ perf#rmane $ata'
 A. rinciple
Te*t *ample i* extrate$ in <arm et"an#l +*in. refl+x #n$en*er' Extrat i* filtere$
an$ in;ete$ int# li-+i$ "r#mat#.rap" e-+ippe$ <it" U? #r fl+#re*ene $etet#r'
 B. Apparatus 7a8 Li-+i$ "r#mat#.rap" 7LC8'44E-+ippe$ <it" I ? inte.rat#r9 B( .L
*ample in;et#r9 an$ <it" U? $etet#r *et at B( r+n' <a,elen.t" #r fl+#r#meter
<it" exitati#n B( nm an$ emi**i#n )B& nm' Operatin. #n$iti#n* temperat+re9
am2ient 7B(4B&C8@ fl#< rate9 0'& mLmin'9 i*#rati@ relati,e retenti#n time* N,anill:l4
n4n#nanami$e9 0'((@ n#r$i":$r#ap*aiin9 ('%(@ ap*aiin9 0'((@
$i":$r#ap*aiin9 0'&' See /i.+re %%&'() f#r 2a*eline *eparati#n #f ma;#r
ap*aiin#i$*'
728 LC #l+mn'44Stainle**9 C09 0&( x F'5 mm i$9 pae$ <it" & mir#meter partile
*i>e' U*e .+ar$ #l+mn9 if $e*ire$'
78 Refl+x #n$en*er'
7$8 S:rin.e filter'4 ('F& mir#meter'
7e8 C0 *#li$4p"a*e extrati#n artri$.e'
C. !ea"ents
(a# Ethanol 4%&H #r $enat+re$9 *+ita2le f#r "r#mat#.rap":
($# Acetone'4ACS .ra$e'
(c# %C mo$ile p"a*eAet#nitrile4<ater' U*e LC .ra$e *#l,ent*9 #r e-+i,alent' ix
F(( mL aet#nitrile <it" 5(( mL 6B( #ntainin. 0H aeti ai$ 7,,8' De4.a* <it"
"eli+m #r 2: #t"er *+ita2le te"ni-+e'
(d# &'anill)l'n'nonanamide standard solutions. 4N4?anill:ln4n#nanami$e *tan$ar$9
%%H9 i* a,aila2le a* *:nt"eti ap*aiin fr#m Penta Internati#nal C#rp'9 &( Oner
P<:9 Li,in.*t#n9 NJ (3()%' Keep *#l+ti#n* #+t #f $iret *+nli."t' (1# Standard 
solution A'4(' 0& m.mL' A+ratel: <ei." 3& m. N4,anill:l4n4n#nanami$e an$
tran*fer it int# &(( mL ,#l+metri fla*' Dil+te t# ,#l+me <it" et"an#l9 an$ mix'
U*e *tan$ar$ *#l+ti#n A f#r anal:>in. all pepper* exept "ili pepper'7B8  Standard 
solution B.'*.*1+ m"lm%. Tran*fer 0( mL *tan$ar$ *#l+ti#n A int# I (( mL
,#l+metri fla*9 $il+te t# ,#l+me <it" et"an#l9 an$ mix' U*e *tan$ar$ *#l+ti#n 
<"en anal:>in. "ili pepper*'
D' Extrati#n
7a8 ,round or crushed peppers'4A+ratel: <ei." a B& . pepper int# &(( ITIL
2#ilin. fla*' Plae B(( mL et"an#l int# *ame fla*9 a$$ *e,eral .la** 2ea$*9 an$
atta" fla* t# refl+x #n$en*er' Gentl: refl+x te*t *ample & " an$ t"en all#< t#
##l' /ilter 04F mL *ample t"r#+." ('F& .m *:rin.e filter int# *mall .la** ,ial' U*e
f#r LC anal:*i*'
728 !ed pepper oleoresins'4A+ratel: <ei." 04B . #le#re*in int# &( mL ,#l+metri
 

fla*' Inrea*e <ei."t #f *ample9 if t#tal ap*aiin#i$ #nentrati#n i* M 0 H'

N#te D# n#t all#< an: #le#re*in t# #at *i$e* #f fla*'
A$$ & mL aet#ne9 C7289 t# fla* an$ *<irl #ntent* #f fla* +ntil te*t *ample i*
#mpletel: $i*per*e$ 7n# #le#re*in an #at 2#tt#m #f fla* <"en t+rnin. fla* ne 
at F& an.le8' A$$ fi,e )4& mL p#rti#n* et"an#l9 *<irlin. fla* $+rin. ea" a$$iti#n'
Dil+te #ntent* #f fla* t# ,#l+me <it" et"an#l an$ mix <ell'